genetics module 3

Explain how genetic changes can impact biochemical pathways and how phenotypes can result from these changes.

Mechanism: Mutations in genes that encode enzymes create a "block" in a metabolic pathway.

Phenotypic Result: The final end product is not formed, leading to a mutant phenotype.

Intermediate accumulation

The compound immediately preceding the blocked step accumulates in the cell.

examples of biochemical pathway blockage

• Alkaptonuria: absence of homogentisate oxidase activity, leading to a build-up of homogentisic acid

• Pku: an accumulation of phenylpyruvic acid in the brain due to a block in phenylalanine hydroxylase

• Albinism: albinism block limits production of melanin

Explain how biochemical pathways can have epistatic interactions.

a mutation in the precursor prevent the intermediate from forming, disregarding the formation of the second step

survival rule

A mutant strain will only grow if provided with a compound created after the blocked step.

how did early scientists determine biochemical pathways

used neospora: usually it can synthesize arginine on its own, but mutants need supplements to do this

ordering compounds in biochemical pathway

The compound that supports growth in the most mutants is likely the final product; the compound that supports growth in the fewest is early in the pathway.

one gene = ____

one gene = one polypeptide/enzyme rule

Each step in a pathway is typically catalyzed by an enzyme produced by a specific gene.

Explain the significance of mutation in somatic cells versus germline cells.

Somatic: Occur in non-reproductive cells; passed to new cells via mitosis but not to offspring.

Germline: Occur in cells that give rise to gametes; passed to approximately half of the next generation.

Compare and contrast loss of function with gain of function mutations.

Loss of Function: Causes partial or complete absence of normal protein function; usually recessive.

Gain of Function: Produces a protein not normally present or active in a new time/place; usually dominant

silent (synonymous) mutation consequence

Codes for the same amino acid (no change in protein).

neutral mutation consequence

Missense mutation that changes the amino acid but does not affect protein function.

missense (nonsynonymous) mutation consequence

Changes one amino acid to another, potentially altering function

missense mutation example

sickle cell anemia: missense mutation where Glutamate is substituted with Valine (GAG to GUG), which makes the hemoglobin (HbS) unable to bind oxygen as effectively.

nonsense mutation consequence

Changes a sense codon into a stop codon, resulting in a truncated protein.

readthrough mutation consequence

Changes a stop codon into a sense codon, resulting in a longer protein

point mutations

transition or transversion, a single base pair changed

transition

Purine to purine (A↔G) or pyrimidine to pyrimidine (C↔T)

transversion

Purine to pyrimidine (A/G ↔ C/T) or vice versa

frameshift mutation

 insert/deletion of 1 or more base pairs, shifting reading frame (can alter start/stop site)

• 3 insertions/deletion keeps the same chain

depurination

Loss of a purine base (A or G) leaving an apurinic (AP) site and removal of glycosidic bond btwn base and sugar; during replication, any base can be inserted opposite the gap. the base opposite the AP site is unspecified during replication, so the polymerase just "guesses"

deamination

Loss of an amino group; deamination of Cytosine results in Uracil, causing a CG to TA transition.

wobble pairing

Flexibility in the DNA helix allows mispairing (e.g., T with G); requires two rounds of replication to permanently alter both strands.

common causes for spontaneous point mutations

wobble base pairing, depurination, deamination

template slippage

Small loops in the DNA during replication cause the polymerase to add or omit bases

unequal crossing over

Misalignment of homologs during meiosis leads to one chromosome with an insertion and one with a deletion

deamination mutation

C to T/A to G transition

alkylation mutation

G to A transition

hydroxylation

C to T transition

base analogs

causes transition after replication

Chemicals like 5-bromouracil and 2-aminopurine that mimic bases but mispair frequently.

intercalating agents

Proflavin, ethidium bromide, acridine orange

• Large, planar molecules that slip between base pairs of DNA. This distorts the

helix, causing template slippage during replication.

UV light

Causes pyrimidine (thymine) dimers by forming additional covalent bonds that distort the helix and block replication.

UV light mutation example

Xeroderma pigmentosum – a human disorder where a repair mechanism is defective. Results in tumors on the skin surface

repeat regions

causes hairpin formation and same strand being replicated twice, leading to expansions (more repeats in daughter cell)

repeat region expansion examples

Huntingtons, fragile x syndrome (CGG repeat expansion in the FMR1 gene, which becomes methylated and silenced. more common in males)

x rays

 Breaks chromosomes by breaking phosphodiester bonds. can also cause point mutations. causes most damage when chromosomes are condensed in mitosis

oxidating agents

damages DNA: causes GC to TA transversions

cystic fibrosis

due to mutation in structural protein

Describe how transposons can cause mutations.

DNA sequences that move to new sites; can disrupt genes or regulatory areas and alters phenoytpes

Explain how DS and Ac transposable elements in maize can result in a variegated kernel phenotype.

Ac (Activator) is autonomous; Ds (Dissociation) is non-autonomous and requires Ac's transposase to move. This movement during development creates variegated (spotted) kernel phenotypes.

what purpose do transopons serve

both arabidopsis plants, humans, and drosophila have transopon derived proteins that serve a vital function in their bodies

proofreading

DNA polymerase stalls replication and uses 3’-5’ exonuclease activity to remove incorrect nucleotides immediately after they are added. "trigger": DNA polymerase stalls because the 3’OH is not in the proper position for the next nucleotide

mismatch repari (MMR)

mismatch repair proteins recognize mismatched bases after replication, exonucleases remove new strand from methylated sequence to mismatch, DNA pol fills in the gap; in bacteria, it distinguishes the "old" strand by its methylation.

• CANT remove lesions

direct repair

Corrects damage without replacing the base

direct repair examples

photolyase in bacteria clipping thymine dimers

methyltransferase restoring correct base form to guanine

nucleotide excision repair (NER)

Removes bulky lesions (like thymine dimers) by cutting out a segment of the strand and replacing it.

• In all organisms

• Strands held apart by SSB, enzyme cleaves sugar phosphate bonds on the sides of the lesion, DNA pol fills in gap

base excision repair (BER)

Uses glycosylases to remove specific modified bases (like Uracil), then AP endonuclease removes the sugar-phosphate before replacement. there is one specific glycosylase for each type of modified base (e.g., one for uracil, one for 7-methylguanine)

double strand break repair systems

• Homologous Recombination: Uses the sister chromatid as a template (accurate).uses many of the same enzymes as meiosis

• Non-homologous End Joining (NHEJ): Joins ends directly; prone to errors like deletions or translocations.

translesion dna polymerase

Specialized polymerases that can bypass DNA lesions but are highly error-prone.

deletion

Missing part of a chromosome; creates a compensation loop in heterozygotes.

• Terminal: produces acentric segment which is lost during division

• Interstitial: requires 2 breaks

duplication

Extra copy of a segment; can be tandem (adjacent), displaced (same arm or diff arm), or reverse tandem

• creates compensation loop

• From unequal crossing over

compensation loop

loop of segment that will get deleted in order to align long and short chromosome

• common in tandem duplications

inversion

Segment flipped 180°; requires an inversion loop to synapse in heterozygotes

paracentric inversion

Does not include centromere; produces dicentric bridges (breaks at a random location, and the acentric fragment is lost) and acentric fragments if crossing over occurs.

pericentric inversion

Includes centromere; produces recombinant gametes with duplications/deletions.

translocation

Exchange between nonhomologous chromosomes.

reciprocal translocation

• Two-way exchange of arms, no gain or loss of DNA

  • Example: burkitt’s lymphoma, leads to abnormal b-cell formation

nonreciprocal translocation

segment of one chromosome moved to another, no gain or loss of DNA (segment moves from one chromosome to another without exchange)

robertsonian translocation

Two small (telocentric) chromosomes fuse into one large one, losing small fragments.

• Banding patterns can show derivation relationship between two chromosomes

deletion effects

Pseudodominance: Deletion of a dominant allele allows a recessive allele to be expressed.

Haploinsufficiency: One copy of a gene is not enough for a normal phenotype.

inversion loop effects

• Crossing over still possible

• Paracentric: dicentric bridge breaks at random location, acentric fragments lost

• Pericentric: two of resulting chromatids have too many copies of some genes, and none of others creating nonviable recombinant gametes

• Crossover in heterozygous inversion isn’t viable, but it is viable in homozygotes

Crossing over still occurs within ____, but recombinant products are often ____

inversion loops, inviable

translocation effects

• Reciprocal: ½ of gametes are viable bc ½ are alternate

  • Burkitt’s lymphoma, abnormal function of b cells

• Robersonian: can cause familial down syndrome 

  • Isochromosome: if two chromosome 21s join together

Explain the role of duplications and inversions in the evolutionary process.

Duplications allow "extra" genes to mutate and develop new functions (e.g., hemoglobin vs. myoglobin). Inversions can lead to speciation by preventing gene flow.

structural gene

encodes proteins that are used in metabolism/play structural role in cell

regulatory gene

encodes products that interact w/ other sequences and affects their translation/transcription

regulatory elements

dna sequences that aren’t transcribed but regulate other sequences

eukaryote-specific methods of gene regulation

pre-mRNA splicing, microRNA silencing, transcription in nuc and translation in cytoplasm

Explain how proteins bind DNA

DNA Leucine zipper

• Binding domain, the basic arms bind the major groove of DNA NOT the zipper (zipper used for dimerization)

examples of DNA binding domains

helix turn helix (prokaryotes), leucine zipper, zinc fingers

sequential vs constitutive expression

sequential: cascade of gene expression that turn on in order

constitutive: continuously expressed, always “on”

Describe the differences in negative and positive regulation

Negative: A repressor protein binds to DNA to turn off transcription.

Positive: An activator protein binds to DNA to stimulate transcription

Describe the differences in inducible or repressible regulation

Inducible: Normally OFF; turned ON by a small molecule (inducer).

Repressible: Normally ON; turned OFF by a small molecule (corepressor)

Explain the difference between regulated gene expression and constitutive expression

Regulated genes are expressed only when needed (lac operon in ecoli), while constitutive genes are expressed continuously (like tRNA or rRNA), regardless of conditions

regulatory gene function

lacI, codes for repressor

promoter function

lacP, binds RNA polymerase to allow transcription

operator function

lacO, interacts with repressor

structural genes function

lacZ, lacY, lacA: transcribed and translated into proteins

result of lac operon

polycistronic mRNA transcribed, then translated into three separate products

Illustrate how the lac operon's response to lactose is controlled by negative regulation.

Repressor (lacI) binds the operator when lactose is absent When allolactose (the inducer) is present, it binds the repressor, releasing it from the operator.

Illustrate how the lac operon's response to glucose is controlled by positive regulation

Controlled by CAP and cAMP. Low glucose leads to high cAMP; cAMP-CAP complex binds the promoter to help RNA polymerase

lac operon what happens

lactose gets broken down into galactose and glucose with B-galactosidase

what happens when lactose is absent vs present

absent: operon repressed

present: operon induced

lac operon cis mutation

action of an element only affects genes adjacent to it (ex: operator, promoter)

lac operon trans mutation

diffusible product produced, mutant gene can affect non adjacent genes (ex: repressor)

catabolite repression

glucose inhibits cAMP formation, so no need for lac operon function

not transcribing operons that aren’t needed

what is required for activation of lac operon

CAP protein (so that camp can bind cap)

glucose ____ causes low cAMP

import

Illustrate how the trp operon's response to tryptophan is controlled by negative regulation.

Repressor only binds the operator when tryptophan (the corepressor) is present bc there is less transcription when its present

Explain regulation by attenuation in the trp operon

Using the movement of the ribosome to determine whether transcription should finish.

Ribosome begins translating the mRNA leader sequence while RNA polymerase is still transcribing it.

• If tryptophan levels are low, the ribosome stalls at two consecutive Trp codons, allowing the polymerase to get ahead and enabling regions 2 and 3 of the mRNA to base-pair; this "anti-terminator" structure allows transcription to continue so the cell can make more tryptophan

• If tryptophan is present, the ribosome does not stall and quickly moves to block region 2, which forces regions 3 and 4 to pair; this creates a hairpin structure that acts as a termination signal, causing RNA polymerase to fall off before the structural genes are even reached.

explain why attenuation is restricted to prokaryotes.

bc prokaryotes dont have as much compartmentalization as eukaryotes, so they can do transcription and translation simultaneously

trp leader sequence

usually 5’ has an untranslated region, but trp is translated to create regulatory sequence

anti sense rna how does it post-transcription regulate

• Mechanism: Small RNA molecules produced from a different gene that are complementary to a specific mRNA sequence.

• Action: They base pair with the mRNA, physically blocking the ribosome from binding or moving, thereby inhibiting translation.

Example: In E. coli, adjusting ompF and ompC membrane proteins allows adaptation to different ion concentrations.

riboswitches how does it post-transcription regulate

• Definition: Specific RNA sequences located within the mRNA molecule itself (typically the 5' UTR).

• Mechanism: When a specific effector molecule binds to the riboswitch, the mRNA undergoes a conformational change.

Action: This change can hide or expose the ribosome-binding site, directly regulating whether the mRNA is translated

List and define the levels of regulation of gene expression in eukaryotes.

Changes in Chromatin: Histone modification, chromatin remodeling, and DNA methylation.
+1

Initiation of Transcription: Involves transcription factors, activators, repressors, and insulators.
+1

RNA Processing and Stability: RNA splicing, degradation, and interference.
+1

Protein Modification: Post-translational changes to the final protein product

Defend how most cells can have the same genetic content and yet have different functions in the body.

Cellular Differentiation:

1. Concept: Almost all cells in a multicellular organism contain the same genome (genetic content).

Defense: Differences in function and structure arise from differential gene expression—turning specific sets of genes on or off at different times and in different cell types

Compare the transcriptional potential of condensed chromatin, decondensed chromatin, and naked DNA.

Naked DNA: Highest potential; no physical barriers to RNA polymerase.

Decondensed Chromatin (11nm fiber): High potential; "loosely coiled" and DNaseI sensitive sites, allowing regulatory proteins to access binding sites.

Condensed Chromatin: Low/No potential; tightly coiled DNA is physically inaccessible to the transcription machiner

how does histone acetylation affect gene expression

HAT enzymes add acetyl groups to histone tails, neutralizing their positive charge and loosening the DNA-histone bond to promote transcription.

how does histone deacetylation affect gene expression

HDAC enzymes remove acetyl groups, tightening the DNA-histone association and repressing transcription.

Describe what DNA methylation is and how it affects gene expression.

Addition of methyl groups to cytosine bases (often in CpG islands); typically leads to gene silencing by attracting deacetylases or physically blocking transcription factors